小麦抗穗发芽研究进展

穗发芽严重影响小麦品质和产量。种子自身休眠特性、α-淀粉酶活性、α-淀粉酶抑制剂、迟熟α-淀粉酶活性、种皮颜色、颖壳抑制物以及穗部形态等,均是影响小麦穗发芽的重要因素,其中对子粒休眠特性和α-淀粉酶活性的研究较为深入。位于第3染色体组上的R基因、休眠基因以及4AL上的Phs基因均与小麦穗发芽密切相关。已开发出一些与穗发芽抗性相关的分子标记,其中位于第3部分同源群的三重R基因和位于3B染色体的STS标记Vp1B3,以及位于3A染色体的主效QTL位点QPhs.ccsu-3A.1均可直接用于穗发芽抗性的筛选。本文对以上内容进行了详细论述,并就今后如何提高小麦穗发芽抗性进行了讨论。     

小麦穗发芽鉴定方法的比较与分析

穗发芽是小麦生产中较为严重的灾害之一,易受外界环境的影响,一旦发生不仅会影响产量,而且还会严重影响小麦的品质,因此培育抗穗发芽的小麦品种至关重要。该研究通过对65份小麦材料进行穗发芽试验,比较分析了小麦穗发芽抗性的常用方法,即籽粒发芽法、整穗发芽法和大田穗发芽法。结果表明:三种方法之间均呈极显著正相关关系,而且在1%水平上均存在极显著性差异;发芽指数与籽粒发芽率的相关性最高,能够更好地评价小麦材料的休眠特性,但不能得出材料的总体抗性;籽粒发芽法和整穗发芽法的变异程度相对较小,试验条件更易控制,可作为小麦穗发芽抗性评价的简易方法;多数参试材料的平均籽粒发芽率平均整穗发芽率平均大田穗发芽率,且三者差异程度均达到极显著水平,这说明麦穗的外部结构及外部环境对小麦穗发芽的影响显著。因此,籽粒发芽法可以从休眠性方面,对小麦种子资源进行初步筛选;整穗发芽法可用于穗发芽抗性的进一步鉴定和验证,评价小麦材料穗发芽的综合抗性;大田穗发芽法较易受自然条件的影响、变异程度较大,其结果可以作为室内发芽试验的参考数据。     

RSP抗穗发芽基因育种利用研究初报

利用农艺性状优良、优质的红粒小麦品种绵阳11以及2个白粒、极易穗发芽(常年穗发芽在50%以上)的小麦品系YY2和88-1643与穗发芽抗性来源于长休眠节节麦的人工合成小麦RSP杂交,对3个组合的F2单株和F3株系的穗发芽测试,从RSP×绵阳11中筛选出15个与RSP相当的纯合抗性株系和27份穗发芽接近0的F3单株;从RSP×YY2和RSP×88-1643组合中共筛选出5份穗发芽在7%以下的白粒株系.这些株系或单株相对于RSP的高秆、晚熟等不利性状已有较大改良,为节节麦抗穗发芽基因向优质或白粒小麦转移研制出了更容易利用的中间材料.     

珍稀植物青檀种子休眠与萌发的研究

为了探讨和研究青檀种子休眠和萌发特性,采用石蜡切片法、生物鉴定(白菜籽发芽实验)法、赤霉素溶液浸种、以及赤霉素与低温层积相结合等方法,寻找引起青檀种子休眠的原因和解除休眠的最佳措施。结果表明:青檀种子本身含有发芽抑制物和存在生理后熟是引起休眠的2个主要原因,用质量浓度为300mg/L的赤霉素溶液浸种24h或低温层积后用赤霉素处理均能在一定程度上解除休眠促进萌发,其中以低温层积25d后用500mg/L的赤霉素浸种36h效果最好。发芽率和发芽势分别达到83.5%和65%。     

小麦Vp-1基因RNA干扰表达载体的构建及遗传转化

小麦成熟期穗发芽是世界性的自然灾害,严重影响小麦的产量和品质.Viviparous-1(Vp-1)是促进胚成熟和休眠的主要转录调节因子,与小麦穗发芽抗性有着密切的关系.本实验根据小麦Vp-1基因序列,以植物表达载体pAHC25为基础,成功构建了含有反向重复序列的RNA干扰表达载体pAHC-WVpRi.采用基因枪法轰击小麦品种新春9号幼胚材料1825个,共获得34株T0再生植株.利用Bar基因引物和干扰片段特异引物对再生植株进行PCR检测,获得Bar基因和干扰片段均为阳性的植株3株,转化率为0.16%.本研究为深入分析Vp-1基因功能,进而通过分子育种进行小麦穗发芽抗性的遗传改良提供了科学依据.     

不同种源的葡萄种子休眠及其解除的研究

为探讨葡萄种子休眠与解除的规律, 我们选择起源于东亚、北美–中美、欧洲–中亚3个分布中心的不同种类及其杂种的20个品种, 研究了它们成熟种子的外部形态与萌发行为, 种子的休眠特性与休眠解除的方法, 并模拟四季温度的交替变化研究了环境温度对种子休眠的影响。结果表明, 不同起源的各类葡萄种子的休眠类型均为生理休眠, 但其休眠程度不同, 休眠解除方式也存在差异。其中欧亚种和东亚种的种子休眠较浅, 美洲种种子休眠较深; 杂交种比亲本所属类别的种子休眠程度浅。对于欧亚种、东亚种及其杂交种(欧山杂种)而言, 5ºC冷层积和变温层积(即20ºC (14 h) /10ºC (10 h)和30ºC (14 h) /20ºC (10 h))2个月能够有效地或部分解除它们的种子休眠; 但对美洲种和欧美杂种而言, 仅5ºC冷层积且层积时间需要延长至6个月才能解除其休眠, 变温层积和25ºC暖层积都不能解除休眠。四季温度的交替变化模拟实验进一步证明了不同起源的葡萄种子的休眠程度不同。这些休眠特性及其解除方式反映了不同起源葡萄种类的环境适应性。本文研究结果为葡萄资源的引种和育种提供了参考数据。     

利用RIL和CSSL群体检测水稻种子休眠性QTL

利用由梗稻品种Asominori与籼稻品种IR24的杂交组合衍生的重组自交F10。家系(Recombinant Inbred Lines,RIL)群体及其衍生的染色体片段置换系(Chromosome Segment Substitution Lines,CSSL)群体,进行了种子休眠性QTL的检测和遗传效应分析。其中CSSL群体有2个,即CSSLl(以Asominori为背景,置换片段来自IR24)和CSSL2(以IR24为背景,置换片段来自Asominori)。在RIL群体上共检测到3个种子休眠性QTL,分别位于第3、6和9染色体上;在CSSL1群体中检测到分布在第1、3和7染色体上的3个休眠性QTL;而在CSSl2群体上检测到的3个QTL则分别位于第1、2和7染色体上。同时在两套CSSL群体上,分别检测到位于第1、7染色体上位置相近且效应一致的休眠性QTL,分析表明其所在的Asominori片段含对种子休眠性的增效基因,相应的IB24段含有减效基因。     

豆科植物种子物理性休眠解除机制的研究进展

物理性休眠普遍存在于豆科植物种子中,其休眠解除机制的研究对阐明自然条件下种群的繁殖更新、生态环境恢复及种质资源的保存等都具有重要意义.本文对国内外关于影响种子休眠解除的环境因素(温度与温度的波动、湿度或土壤含水量)和生物因素(动物捕食、微生物感染等),休眠解除的结构基础,以及各种因子作用于种子的方式和部位的研究进展进行综述,并简要评述了休眠解除机制的二阶段和四阶段假说,以期为豆科植物种子休眠的进一步研究提供参考.     

温度对青藏高原高寒草地植物种子萌发与病原真菌侵染关系的影响

自然条件下,病原真菌侵染是导致种子死亡的重要因素。气候变化背景下,种子特性(如休眠和发芽速率)与真菌侵染之间的关系仍不清楚。本研究以青藏高原高寒草地的17种草本植物种子为对象,设置不同温度(10、15、20和25℃)发芽实验,研究不同温度条件下种子病原真菌侵染和死亡与种子发芽速率和休眠特性之间的关系。结果表明：在各温度条件下,种子病原真菌侵染率、死亡率与种子休眠率均呈显著负相关(P<0.001),但在25℃条件下相关性最强：仅在25℃条件下,种子真菌侵染率、死亡率与种子平均发芽时间呈显著正相关(P<0.05);种子休眠在不同温度条件下均能有效抵御病原真菌的侵染;种子通过快速萌发有效逃避病原真菌的侵染,但其与病原真菌的关系受温度影响较大。     

GA3和变温层积对天女木兰种子萌发及内源激素的影响

用不同质量浓度GA3浸泡天女木兰种子并结合变温层积处理,应用高效液相色谱法对不同时期种子中4种激素GA3、IAA、ABA、ZR含量进行测定,并测量种胚长和萌发率,以探讨天女木兰种胚发育,内源激素含量变化与种子休眠萌发之间的调控关系,为进一步研究种子休眠机理提供理论基础。结果表明:(1)天女木兰成熟种子胚发育不完全,胚乳内高浓度ABA和低浓度GA3是其休眠的主要原因。(2)GA3处理能促使天女木兰种子提前30d完成形态后熟,并以1 500mg·L-1 GA3处理效果最佳。(3)在变温层积过程,天女木兰种胚发育可分三个阶段:阶段Ⅰ(0~70d)完成种胚进一步分化;阶段Ⅱ(70~120d)种胚快速生长时期;阶段Ⅲ(120~150d)休眠完全解除,种子具备发芽能力。种子能否打破休眠主要取决于阶段Ⅰ和Ⅱ的状况。(4)GA3/ABA、IAA/ABA和ZR/ABA在种子后熟期间的变化同胚生长发育存在一致性,认为内源激素的相对水平对种子休眠具有重要的调控作用。     

MPK9 and MPK12 function in SA-induced stomatal closure in Arabidopsis thaliana

Salicylic acid (SA) induces stomatal closure sharing several components with abscisic acid (ABA) and methyl jasmonate (MeJA) signaling. We have previously shown that two guard cell-preferential mitogen-activated protein kinases (MAPKs), MPK9 and MPK12, positively regulate ABA signaling and MeJA signaling in Arabidopsis thaliana. In this study, we examined whether these two MAPKs are involved in SA-induced stomatal closure using genetic mutants and a pharmacological, MAPKK inhibitor. Salicylic acid induced stomatal closure in mpk9 and mpk12 single mutants but not in mpk9 mpk12 double mutants. The MAPKK inhibitor PD98059 inhibited SA-induced stomatal closure in wild-type plants. Salicylic acid induced extracellular reactive oxygen species (ROS) production, intracellular ROS accumulation, and cytosolic alkalization in the mpk9, mpk12, and mpk9 mpk12 mutants. Moreover, SA-activated S-type anion channels in guard cells of wild-type plants but not in guard cells of mpk9 mpk12 double mutants. These results imply that MPK9 and MPK12 are positive regulators of SA signaling in Arabidopsis guard cells.     

Cadmium activates Arabidopsis MPK3 and MPK6 via accumulation of reactive oxygen species

Cadmium (Cd) is a non-essential toxic heavy metal that influences normal growth and development of plants. However, the molecular mechanisms by which plants recognize and respond to Cd remain poorly understood. We show that, in Arabidopsis, Cd activates the mitogen-activated protein kinases, MPK3 and MPK6, in a dose-dependent manner. Following treatment with Cd, these two MAPKs exhibited much higher activity in the roots than in the leaves, and pre-treatment with the reactive oxygen species (ROS) scavenger, glutathione, effectively inhibited their activation. These results suggest that the Cd sensing signaling pathway uses a build-up of ROS to trigger activation of Arabidopsis MPK3 and MPK6.     

Two Arabidopsis guard cell-preferential MAPK genes,MPK9 and MPK12, function in biotic stress response

Abscisic acid (ABA) plays a major role in plant development and adaptation to severe environmental conditions. ABA evokes cellular events to regulate stomatal apertures and thus contributes to the plant’s ability to respond to abiotic stresses. Reactive oxygen species (ROS) are produced in response to ABA and mediate ABA-induced stomatal closure. We have shown that two MAP kinases, MPK9 and MPK12, are highly and preferentially expressed in guard cells and function as positive regulators of ROS-mediated ABA signaling in guard cells. Cell biological and electrophysiological analyses demonstrated that MPK9 and MPK12 act downstream of ROS and cytosolic Ca²⁺ and upstream of anion channels in the guard cell ABA signaling cascade. Plant pathogens use stomata as the primary gateway to enter into their hosts, and previous studies have indicated crosstalk between ABA and defense signaling. Here we show that mpk9-1/12-1 double mutants are highly susceptible to Pseudomonas syringae DC3000 compared to WT plants. These results suggest that the regulation of stomatal apertures by MPK9 and MPK12 contributes to the first line of defense against pathogens.     

MAPK phosphatase MKP2 mediates disease responses in Arabidopsis and functionally interacts with MPK3 and MPK6

Mitogen‐activated protein kinase (MAPK) cascades have important functions in plant stress responses and development and are key players in reactive oxygen species (ROS) signalling and in innate immunity. In Arabidopsis, the transmission of ROS and pathogen signalling by MAPKs involves the coordinated activation of MPK6 and MPK3; however, the specificity of their negative regulation by phosphatases is not fully known. Here, we present genetic analyses showing that MAPK phosphatase 2 (MKP2) regulates oxidative stress and pathogen defence responses and functionally interacts with MPK3 and MPK6. We show that plants lacking a functional MKP2 gene exhibit delayed wilting symptoms in response to Ralstonia solanacearum and, by contrast, acceleration of disease progression during Botrytis cinerea infection, suggesting that this phosphatase plays differential functions in biotrophic versus necrotrophic pathogen‐induced responses. MKP2 function appears to be linked to MPK3 and MPK6 regulation, as indicated by BiFC experiments showing that MKP2 associates with MPK3 and MPK6 in vivo and that in response to fungal elicitors MKP2 exerts differential affinity versus both kinases. We also found that MKP2 interacts with MPK6 in HR‐like responses triggered by fungal elicitors, suggesting that MPK3 and MPK6 are subject to differential regulation by MKP2 in this process. We propose that MKP2 is a key regulator of MPK3 and MPK6 networks controlling both abiotic and specific pathogen responses in plants.     

Determination of primary sequence specificity of Arabidopsis MAPKs MPK3 and MPK6 leads to identification of new substrates

MAPKs (mitogen-activated protein kinases) are signalling components highly conserved among eukaryotes. Their diverse biological functions include cellular differentiation and responses to different extracellular stress stimuli. Although some substrates of MAPKs have been identified in plants, no information is available about whether amino acids in the primary sequence other than proline-directed phosphorylation (pS-P) contribute to kinase specificity towards substrates. In the present study, we used a random positional peptide library to search for consensus phosphorylation sequences for Arabidopsis MAPKs MPK3 and MPK6. These experiments indicated a preference towards the sequence L/P-P/X-S-P-R/K for both kinases. After bioinformatic processing, a number of novel candidate MAPK substrates were predicted and subsequently confirmed by in vitro kinase assays using bacterially expressed native Arabidopsis proteins as substrates. MPK3 and MPK6 phosphorylated all proteins tested more efficiently than did another MAPK, MPK4. These results indicate that the amino acid residues in the primary sequence surrounding the phosphorylation site of Arabidopsis MAPK substrates can contribute to MAPK specificity. Further characterization of one of these new substrates confirmed that At1g80180.1 was phosphorylated in planta in a MAPK-dependent manner. Phenotypic analyses of Arabidopsis expressing phosphorylation site mutant forms of At1g80180.1 showed clustered stomata and higher stomatal index in cotyledons expressing the phosphomimetic form of At1g80180.1, providing a link between this new MAPK substrate and the defined role for MPK3 and MPK6?in stomatal patterning.     

Effects of MPK3 and MPK6 kinases on the chloroplast architecture and function induced by cold acclimation in Arabidopsis

Photosynthesis Research - Exposure to low, non-freezing temperatures develops freezing tolerance in many plant species. Such process is called cold acclimation. Molecular changes undergone during...     

Arabidopsis MPK3 and MPK6 play different roles in basal and oligogalacturonide- or flagellin-induced resistance against Botrytis cinerea

Mitogen-activated protein kinases (MAPKs) are fundamental components of the plant innate immune system. MPK3 and MPK6 are Arabidopsis (Arabidopsis thaliana) MAPKs activated by pathogens and elicitors such as oligogalacturonides (OGs), which function as damage-associated molecular patterns, and flg22, a well-known microbe-associated molecular pattern. However, the specific contribution of MPK3 and MPK6 to the regulation of elicitor-induced defense responses is not completely defined. In this work we have investigated the roles played by these MAPKs in elicitor-induced resistance against the fungal pathogen Botrytis cinerea. Analysis of single mapk mutants revealed that lack of MPK3 increases basal susceptibility to the fungus, as previously reported, but does not significantly affect elicitor-induced resistance. Instead, lack of MPK6 has no effect on basal resistance but suppresses OG- and flg22-induced resistance to B. cinerea. Overexpression of the AP2C1 phosphatase leads to impaired OG- and flg22-induced phosphorylation of both MPK3 and MPK6, and to phenotypes that recapitulate those of the single mapk mutants. These data indicate that OG- and flg22-induced defense responses effective against B. cinerea are mainly dependent on MAPKs, with a greater contribution of MPK6.     

Haplo-insufficiency of MPK3 in MPK6 mutant background uncovers a novel function of these two MAPKs in Arabidopsis ovule development

The plant life cycle includes diploid sporophytic and haploid gametophytic generations. Female gametophytes (embryo sacs) in higher plants are embedded in specialized sporophytic structures (ovules). Here, we report that two closely related mitogen-activated protein kinases in Arabidopsis thaliana, MPK3 and MPK6, share a novel function in ovule development: in the MPK6 mutant background, MPK3 is haplo-insufficient, giving female sterility when heterozygous. By contrast, in the MPK3 mutant background, MPK6 does not show haplo-insufficiency. Using wounding treatment, we discovered gene dosage-dependent activation of MPK3 and MPK6. In addition, MPK6 activation is enhanced when MPK3 is null, which may help explain why mpk3(-/-) mpk6(+/-) plants are fertile. Genetic analysis revealed that the female sterility of mpk3(+/-) mpk6(-/-) plants is a sporophytic effect. In mpk3(+/-) mpk6(-/-) mutant plants, megasporogenesis and megagametogenesis are normal and the female gametophyte identity is correctly established. Further analysis demonstrates that the mpk3(+/-) mpk6(-/-) ovules have abnormal integument development with arrested cell divisions at later stages. The mutant integuments fail to accommodate the developing embryo sac, resulting in the embryo sacs being physically restricted and female reproductive failure. Our results highlight an essential function of MPK3 and MPK6 in promoting cell division in the integument specifically during ovule development.